The present invention relates to a cellulosic particle body, a method of producing said particle body, a spherical type body which comprises crosslinked polymer particles interconnected with the aid of an organic binder comprising a non-crosslinked polymer, a method of producing said spherical type body, and an adsorbent for purification of body fluids which is capable of removing a target substance at a high speed in the therapy of hyperlipemia, autoimmune diseases and immunity-mediated diseases and the like.
A cellulosic particle body and a spherical type body comprising a crosslinked polymer particle are in broad use in a variety of fields, for example as a support for immobilization of microbial cells or enzymes, an adsorbent matrix for perfumes and pharmaceuticals, an adsorbent for purification of body fluids, a cosmetic additive, a chromatographic stationary phase material, etc. or, thorough introduction of a functional group, even as various ion exchangers.
Much research has been undertaken into the cellulosic particle body.
Japanese Kokai Publication Sho-63-90501 discloses a technology which comprises blending an anionic water-soluble compound with a mixture of viscose and a water-soluble macromolecular compound to prepare a dispersion of microfine particles, coagulating the dispersion under heating or with the aid of a coagulant, regenerating it with an acid, and removing the water-soluble macromolecular compound through a series of coagulation, regeneration and aqueous washing to provide a porous microfine cellulosic particle body with a mean particle diameter of not greater than 3xc3x9710xe2x88x924 m and a maximum pore volume within a pore volume range of 2xc3x9710xe2x88x928 to 8xc3x9710xe2x88x927 m, with the total pore volume of all the pores within said range being not less than 2.5xc3x9710xe2x88x925 m3/kg. The particle body provided by the above technology is such that the cellulosic particle body as such have fine pores.
Japanese Kokai Publication Sho-63-92602 discloses a technology which comprises blending viscose, calcium carbonate and a water-soluble anionic macromolecular compound to prepare a dispersion of finely divided particles of calcium carbonate-containing viscose, coagulating and neutralizing the dispersion, and decomposing calcium carbonate with an acid to provide a porous cellulosic particle body.
With those technologies, however, the cellulosic particle body obtained are relatively small in diameter, so that in certain applications such as a filler, an adsorbent, etc., it is difficult to carry out a large-scale treatment at a high flow rate and if a high-speed treatment is attempted, the cellulosic particle body tend to be destroyed. Moreover, when such a cellulosic particle body is used for the treatment of body fluids, plugging with blood corpuscles tend to take place.
Accordingly there has been a demand for development of a cellulosic particle body which would have sufficiently high mechanical strength, be compatible with treatment at high flow rates, exploit the pore structure of the cellulosic particle body providing for a large surface area, and be free from the trouble of plugging in the treatment of body fluids.
Meanwhile, in the field of body fluid treatment, a body fluid purification method is being practiced as a therapeutic technique comprising removal of a specific substance(s) from a body fluid, which comprises passing the body fluid through an adsorption device packed with an adsorbent immobilized a substance having an affinity for a target substance on a carrier to thereby adsorb and remove said substance. The method developed initially for this purpose comprised passing whole blood over active charcoal, particularly a coated charcoal particle to remove a target substance. With advances in plasma perfusion membranes, various adsorption devices for removing a target substance from separated plasma have been developed.
Generally speaking, in body fluid purification therapy, the treatment time is preferably as short as possible from the standpoint of the patient""s quality of life. For reducing the treatment time, several approaches may be contemplated by using ingenuity in the aspect of operating conditions with the adsorbent material being held unchanged.
First, it may be contemplated to increase the flow rate of the body fluid in the extracorporeal circuit so as to increase the volume of the body fluid to be contacted with an adsorbent per unit time. However, it will adversely affect the patient""s quality of life to excessively increase the flow rate of the body fluid withdrawn from the patient""s body and circulated extracorporeally. The conventional flow rate of a body fluid for extracorporeal circulation is 0.833xc3x9710xe2x88x926 to 3.33xc3x9710xe2x88x926 m3/s (50 to 200 ml/min.). Thus, there is a limit to the flow rate of the body fluid which can be circulated extracorporeally.
It may also be contemplated to increase the capacity of the adsorption apparatus and thereby prolong the time of contact between the body fluid and the adsorbent. However, as the device capacity is increased, the volume of the body fluid existing outside the body during treatment is increased to adversely affect the patient""s quality of life, with the result that the device capacity cannot be increased beyond a certain limit. The capacity of the conventional adsorption apparatus for purification of a body fluid is 50xc3x9710xe2x88x926 to 500xc3x9710xe2x88x926 m3 (50 to 500 ml) at most.
Then it may also be contemplated to reduce the treatment time by increasing the static adsorptivity of the adsorption apparatus. The static adsorptivity means the saturated amount of adsorption. As a means for enhancing the static adsorptivity, it may be contemplated to enhance the static adsorptivity by increasing the amount of adsorption per unit adsorbent. The factors influencing the adsorption equilibrium relation are the substance having an affinity for the target substance and the contact area effective for adsorbing the target substance. However, said substance having an affinity for the target substance is restricted to a substance having a specific affinity for the particular target substance. Furthermore, it is restricted to a substance substantially not affecting the patient""s physiology because the objective is the treatment of a body fluid. It may also be contemplated to increase the effective contact area but, as the minimum requirement, this contact area must have pores receptive to the target substance. Therefore, the maximum contact area of the porous body having such pores is limited by the diameter and number of pores. Thus, there is a limit to enhancing the static adsorptivity by improving the above-mentioned adsorption equilibrium relation.
As mentioned above, because of the restrictions associated with the body fluid purification technology, it has been found difficult to reduce the treatment time, with the amount of adsorption maintained, by improving the device capacity, the flow rate of a body fluid, and said static adsorptivity.
Lastly, it may be contemplated to reduce the treatment time by improving the dynamic adsorptivity of the adsorption apparatus. The dynamic adsorptivity means the magnitude of adsorption rate. As a means for improving the dynamic adsorptivity, it may be contemplated, for instance, to improve the dynamic adsorptivity by optimizing the particle diameter of the adsorbent and the intraparticle diffusion coefficient of the target substance.
Referring to the first approach, i.e. the method of reducing the particle diameter of the adsorbent and, hence, said diffusion distance to thereby improve the dynamic adsorptivity, reducing the particle diameter of the adsorbent results in a reduced diameter of the fluid flow passageway and an increased pressure loss so that the risk for plugging is increased. Therefore, in consideration of the safety of therapy, the particle diameter cannot be reduced too much. Actually, the particle diameter of the conventional adsorbent for plasma perfusion is 50xc3x9710xe2x88x926m to less than 1000xc3x9710xe2x88x926 m and that for direct blood perfusion is 100xc3x9710xe2x88x926 m to less than 4000xc3x9710xe2x88x926 m.
Referring to the second approach, that is the method which comprises increasing the diffusion coefficient of the target substance within the adsorbent for insuring a fast transfer of the target substance within the adsorbent to hereby improve the dynamic adsorptivity, this method is also subject to the following restrictions. In the case of the conventional adsorbent for purification of a body fluid which depends on rate-determining diffusion, once the target substance is established, its diffusion coefficient has a constant value according to the structure of the adsorbent so that it becomes necessary to add ingenuity to the adsorbent structure. However, even if the structure is optimized, the diffusion coefficient of the target substance within the adsorbent does not increase beyond the diffusion coefficient in the body fluid where no steric hindrance exists and, therefore, this method is also limited.
Thus, as far as the conventional adsorbent for purification of a body fluid is concerned, there is a limit to improving the dynamic adsorptivity by increasing the particle diameter of the adsorbent and the intraparticle diffusion coefficient of the target substance, with the result that the treatment time can hardly be reduced.
On the other hand, while it is difficult to apply them to the purification of a body fluid, there exists some adsorbent materials which, when used as chromatographic carriers for immobilization of a substance having an affinity for the target substance, can be expected to achieve an improved dynamic adsorptivity.
The principles relating to dynamic adsorptivity are now explained in the first place. As an indicator of dynamic adsorptivity, it is common practice to use a breakthrough curve which represents the time course of change in the concentration of the target substance at the exit of an adsorption apparatus when a solution containing said target substance in a given concentration is passed at a constant flow rate. In estimating the dynamic adsorptivity of an adsorption apparatus under operating conditions, it is preferable to keep the linear velocity of flow within the adsorption apparatus constant, that is to say a constant state of flow around the adsorbent. It should be noted that the term xe2x80x9clinear velocity within the adsorption apparatusxe2x80x9d is used in this specification to mean the rate of transfer (m/s) of the mobile phase in the adsorption apparatus.
On the other hand, the theoretical plate number is generally used as an indicator of the performance of a column packed with an adsorbent not carrying an adsorbate thereon (a packed column). The theoretical plate number means the minimum multiples of column height which would be required for a target substance to attain an adsorption-desorption equilibrium when a solution containing it is passed through the packed column.
According to Kato et al. [Shigeo Kato, et el., Journal of Fermentation and Bioengineering, 78, 246 (1994)], the above-mentioned breakthrough curve representing the dynamic adsorptivity of an adsorption apparatus can be correlated with the above-mentioned theoretical plate number as an indicator of the performance of a packed column by the following three expressions.             C              C        0              =          1      -                        ⅇ                                    -              N                        ⁢                          xe2x80x83                        ⁢            θ                          ⁢                  {                      1            +                          N              ⁢                              xe2x80x83                            ⁢              θ                        +                                                            (                                      N                    ⁢                                          xe2x80x83                                        ⁢                    θ                                    )                                2                                            2                !                                      +                          xe2x80x83                        ⁢            …            ⁢                          xe2x80x83                        +                                                            (                                      N                    ⁢                                          xe2x80x83                                        ⁢                    θ                                    )                                                  N                  -                  1                                                                              (                                      N                    -                    1                                    )                                !                                              }                                t      -        =                  α        ⁢                  xe2x80x83                ⁢        V            F            α    =                  q        0                    C        0            
wherein t represents time (in seconds); C represents the concentration [kg/m3] of a target substance at the exit of an adsorption apparatus, which is a time-dependent variable; C0 represents the concentration [kg/m3] of the target substance entering the adsorption apparatus, which is a constant; V represents the volume of the adsorption apparatus or the volume of a packed column [m3], which is a constant; q0 represents the amount of adsorption at equilibrium [kg/m3] at C0, i.e. the amount of adsorption which does not increase any further when a solution of the concentration C0 is passed through the adsorption apparatus, which is a constant; F represents the flow rate [m3/sec.] of solution selected so as to be equal to the linear velocity within the adsorption apparatus under operating conditions, which is a constant; N represents the theoretical plate number as found for the target substance when a solution containing it is passed through the packed column at the same flow rate F as that found for the same target substance when the same solution is passed through the adsorption apparatus, which is a constant; txe2x88x92 represents the average residence time [seconds] of the target substance in the column; xcex8 represents the percentage of t relative to txe2x88x92; and xcex1 is a parameter representing the adsorption efficiency of an adsorbent.
To demonstrate the influence of the theoretical plate number on the breakthrough curve, suitable values were substituted into the above expressions for calculation. The result is shown in FIG. 1. Referring to FIG. 1, the amount of adsorption per unit volume q [kg/m3]of the absorption apparatus up to each point of time t/txe2x88x92 represents the area above the breakthrough curve, that is the value which can be found by integrating {1-(C/C0)} up to that point of time and dividing the result by the volume of the adsorption apparatus. FIG. 2 shows the time course of the absorption amount q with respect to q0 as calculated by said integration. It can be understood from FIG. 2 that the larger the theoretical plate number of the packed column is, the larger is the adsorption amount which can be adsorbed in a given time and the shorter is the time required for adsorbing a given amount of the substance, indicating that the dynamic adsorptivity of the adsorption apparatus is improved. It is, therefore, clear that the dynamic adsorptivity of an adsorption apparatus can be improved by increasing the theoretical plate number of the packed column.
Furthermore, the theoretical plate number of a packed column is dependent on the minimum column height which is necessary for attaining an adsorption-desorption equilibrium (the height equivalent to a theoretical plate) and the height of the packed column and can be expressed by the following equation.   N  =      L    HETP  
wherein L [m] represents the height of a packed column and HETP [m] represents the height equivalent to a theoretical plate. Since the column height is fixed, increasing the theoretical plate number of the packed column can be attained by reducing the height equivalent to a theoretical plate which is characteristic of the carrier packed, and the dynamic adsorptivity of an adsorption apparatus can be improved by this method. Whereas the theoretical plate number is dependent on the housing geometry and other factors, the height equivalent to a theoretical plate is a characteristic which is solely dependent upon the properties of the adsorbent or solid phase. Stated differently, in discussing the height equivalent to a theoretical plate, it is permissible to use a packed column geometrically different from the adsorption apparatus used for construction of the breakthrough curve, although the linear velocity of flow in the packed column should be equal to that in the adsorption apparatus.
Meanwhile, it is known that when a housing is packed with a particle having flow-through pores extending through each particle and sub-pores communicating with said flow-through pores and smaller in diameter than the flow-through pores as a stationary phase material for chromatography, a stationary phase material for affinity chromatography or a support for immobilization of an enzyme and a solution is passed through the packing at a suitable flow rate, the migration of a solute within the packing is rapid (perfusion effect) compared with the usual particulate adsorbent not having flow-through pores so that the objective operation can be accomplished at a high speed [Japanese Kohyo Publication Hei-4-500726, Japanese Kohyo Publication Hei-6-507313, N. B. Affean et al.: Journal of Chromatography, 519, 1 (1990), Shigeo Kato et al.: Journal of Fermentation and Bioengineering, 78, 246 (1994)]. In this specification, a carrier having a structure such that a flow passing through its particles occurs when there is a flow around said carrier particles and that, when there is a flow of a liquid such as a body fluid around the carrier particles, a portion of the flow passes through the carrier particles owing to the resultant pressure gradient is referred to as a perfusion type carrier. The above-mentioned carrier having flow-through pores and sub-pores is a perfusion type carrier.
The perfusion type carrier is known to be a stationary phase with a smaller height equivalent to a theoretical plate. In other words, because of occurrence of flows passing through the carrier particles, the measured height equivalent to a theoretical plate of such a perfusion type carrier is smaller than that of the conventional carrier in which the mass transfer of the target substance depends solely on diffusion. Therefore, an adsorption apparatus packed with an adsorbent comprising a substance having an affinity for the target substance as immobilized on a perfusion type carrier shows an improved dynamic adsorptivity.
As a typical perfusion type carrier, there is known POROS (trade name), chromatographic carriers available from Perceptive Biosystems (particle diameters 10xc3x9710xe2x88x926 m, 20xc3x9710xe2x88x926 m, 5xc3x9710xe2x88x926m) (Japanese Kohyo Publication Hei-4-500726). However, since those carriers are intended to be used for chromatography, they are available only in small particle diameter ranges in consideration of the ease of packing and flow. Therefore, when a container is packed with this kind of carrier and a fluid from a fermentation tank, a slurry, blood or the like is passed through it, plugging tends to take place owing to the small particle diameter. Moreover, in order to attain a perfusion effect, a solution must be passed at a high linear velocity of not less than 2.8xc3x9710xe2x88x923 m/s.
Heretofore unknown is a perfusion type carrier which is large in particle diameter and provides a perfusion effect even when a solution is passed at a low speed. Neither known to this day is a cellulosic perfusion type carrier. For example, POROS (trade name) mentioned above is a carrier comprising conglomerates of fine particles of a styrene-divinylbenzene copolymer.
On the other hand, porous particles of crosslinked polymers have large specific surface areas and have been used broadly as chromatographic column packings or adsorbents and, furthermore, such particles have been actively developed. Such conglomerates of crosslinked polymer particles may have minute voids between the constituent crosslinked polymer particles of the conglomerate and, therefore, express a variety of functions not obtainable with discrete crosslinked polymer particles. The following technology is available for the construction of spherical type bodies or conglomerates having pores between the adjacent constituent crosslinked polymer particles.
Japanese Kokai Publication Hei-9-25303, for instance, discloses a method for interconnecting particles by way of polymerization which comprises polymerizing a monomer on the surface of crosslinked polymer particles. More particularly, this method comprises dispersing crosslinked polymer particles in a dispersion medium containing a monomer, polyvinyl alcohol, etc. to let the monomer penetrate into the crosslinked polymer particles and then polymerizing the monomer to thereby interconnect the crosslinked polymer particles.
However, because the crosslinked polymer particles are bonded to one another by polymerization, this method requires a complicated polymerization procedure and, moreover, is restricted in the diameter of crosslinked polymer particles which can be bonded together (100xc3x9710xe2x88x926 m at most). In addition, since the monomer so polymerized covers up the entire surface of the crosslinked polymer particles, the inherent functions of the particles are impaired. Another disadvantage is that, after use, the crosslinked polymer particles cannot be reused.
The present invention has for its object to provide a carrier or adsorbent which overcomes the above-mentioned disadvantages.
More particularly, in the light of the above-mentioned arts, it is an object of the present invention to provide a cellulosic particle body which is suited for use in treatments at high flow rates and has excellent mechanical strength and a large surface area and a method of producing said particle body.
In the light of the above-mentioned arts, it is a further object of the invention to provide a cellulosic particle body which can be provided in a relatively large particle diameter range and produces a perfusion effect even when a solution is passed at a comparatively low linear velocity and a method of producing said particle body.
In the light of the above-mentioned arts, it is a still another object of the invention to provide a connected particle body comprising assemblages of crosslinked polymer particles with minute interparticle spaces which (1) can be manufactured by a simpler procedure as compared with the prior art, (2) is less restricted in the available particle diameter of assemblages of crosslinked polymer particles as compared with the prior art, (3) has an exposed area uncovered by a monomer polymerized on the surface of crosslinked polymer particles and consequently allowing expression of the inherent functions of said particles, and (4) permits reusing of the crosslinked polymer particles from the assemblages after use.
It is a still further object of the invention to provide an adsorbent for purification of body fluids which is capable of removing a target substance at a high speed so as to reduce the treatment time with the amount of adsorption maintained at a high level.
The present invention relates to a cellulosic particle body comprising interconnected cellulosic small particles and having voids between particles and to a method of producing said cellulosic particle body which comprises dispersing cellulosic small particles in an alkaline medium and contacting the resulting suspension with a coagulating solution. In this specification, the above technology will be referred to as the first invention.
In another aspect, the present invention relates to a perfusion type cellulosic particle body which comprises porous cellulosic small particles interconnected to have void between the cellulosic small particles as produced by dispersing the porous cellulosic small particles in an alkaline medium to prepare a suspension and contacting the resulting suspension with a coagulating solution and to a method of producing said perfusion type cellulosic particle body which comprises dispersing porous cellulosic small particles in an alkaline medium and contacting the resulting suspension with a coagulating solution. In this specification, this technology will be referred to as the second invention.
The present invention further relates to a spherical type body which comprises crosslinked polymer particles having diameters within a range of 0.1xc3x9710xe2x88x926 to 100xc3x9710xe2x88x923m with a standard deviation of not greater than 100% of their mean diameter and which has a diameter of 1xc3x9710xe2x88x926 m to 100xc3x9710xe2x88x923 m, and satisfies the following conditions (A) to (C):
(A) that said crosslinked polymer particles are interconnected via an organic binder comprising a non-crosslinked polymer;
(B) that the surfaces of said crosslinked polymer particles have area(s) not covered with said organic binder but remaining exposed;
(C) that voids exist between the interconnected crosslinked polymer particles.
The present invention further relates to a method of producing the spherical type body comprising crosslinked polymer particles which comprises immersing crosslinked polymer particles having diameters within a range of 0.1xc3x9710xe2x88x926 m to 10xc3x9710xe2x88x923 m with a standard deviation of not more than 100% of their mean diameter in a solution containing an organic binder comprising a non-crosslinked polymer in an organic solvent which does not dissolve said crosslinked polymer particles but dissolves said organic binder and then evaporating said organic solvent under stirring to interconnect said crosslinked polymer particles via said organic binder separating out on surfaces of said crosslinked polymer particles.
The spherical type body mentioned above need only be substantially spherical and includes a spheroidal body with a ratio of minor axis/major axis up to about 0.7. In this specification, this technology will be referred to as the third invention.
The present invention relates, in a further aspect, to an adsorbent for purification of body fluids which comprises a perfusion type carrier and, as immobilized thereon, a substance having an affinity for a target substance, to an adsorption apparatus for purification of body fluids which comprises a housing packed with said adsorbent and to a method of purifying a body fluid using said adsorption apparatus for purification of body fluids. In this specification, the above technology will be referred to the fourth invention.